Isolation and characterization of the TERE1 gene, a gene down-regulated in transitional cell carcinoma of the bladder.

We have identified a novel cDNA product designated transitional epithelial response gene (TERE1), which was localized to chromosome 1p36. The TERE1 transcript (1.5 and 3.5 kb) is present in most normal human tissues including urothelium, but was reduced or absent in the majority of muscle invasive TCC tumors (22 out of 29 cases). The open reading frame encodes a protein of 338 amino acids (MW 36.8 KD). This protein is 57% homologous to a Drosophila protein called heix. We have shown by Western blotting and immuno-histochemistry with a polyclonal antibody to a specific TERE1 peptide, reduced or absent staining in muscle invasive tumors. Transfection of a sense TERE1 construct resulted in an 80-90% inhibition of cellular proliferation in two TCC cell lines and a lack of aneuploidy in the TERE1-transduced J82 cell line. These data suggest a potential role for this gene product in the progression of bladder cancer.

Influence of breed and ageing time on the sensory meat quality and consumer acceptability in intensively reared beef.

The meat market is very concerned about the influence of ageing on beef quality. However, not many studies have analysed the possible influence of the intrinsic factors (individual, age, sex, body condition, breed, etc.), on the development of the ageing process. The purpose of this study was to assess the influence of breed on the sensory characteristics of the meat throughout the ageing time, using a trained sensory panel and a consumer test. Forty entire males of four breeds, which represented different biotypes (dairy: 10 Spanish Holstein; dual purpose: 10 Brown Swiss; meat type: 10 Limousin; high muscularity: 10 Blonde d'Aquitaine) were analysed. Animals were weaned at seven months on average (except Holstein calves, which were weaned earlier), and fed intensively. Each breed was slaughtered at its usual commercial live weight, according to the market requirements. The m. Longissimus thoracis et lumborum (between T6 and L6) was cut, vacuum packaged and aged for 1, 3, 7, 14, 21 and 35 days. Cooked samples were evaluated by 8 panellists and 200 consumers. Breed had a significant influence on tenderness (p<0.001) and on the quantity of residue after chewing for panellists (p<0.01), but there was a significant interaction between breed and ageing time for tenderness. Long ageing times (more than seven days) tend to reduce differences among breeds in textural characteristics. Ageing had a very important effect on tenderness (p<0.01) and also on some odour and flavour characteristics evaluated by the panel. Taking into account the results obtained in the consumer test, the consumption of the Limousin meat may be recommended at shorter ageing times, meanwhile Blonde d'Aquitaine, Holstein and Old Brown Swiss meats would need a longer ageing period to obtain an optimum acceptance by the consumer.

A murine model of spontaneous aspermatogenesis: linkage to H-2.

Experimental allergic orchitis is an organ-specific autoimmune disease characterized by inflammatory infiltrates associated with the seminiferous tubules of the testes. Orchitis is often, but not always, accompanied by aspermatogenesis in susceptible strains of mice. In this study, various strains of H-2 congenic mice were used to examine the relationship between orchitis and aspermatogenesis, and as a result, a genetic predisposition to spontaneous aspermatogenesis has been defined. A high correlation was seen between orchitis and aspermatogenesis in B10.D2/nSnJ mice, however, the two conditions were uncorrelated in C57BL/10J mice. Subsequent analysis of C57BL/10J congenic strains showed their aspermatogenesis to be spontaneous, rather than due to either testis-specific antigen or adjuvants. Further studies using other H-2 congenic strains revealed that the aspermatogenesis seen in C57BL/10J mice is linked to H-2 and influenced by C57BL/10J background genes. Finally, spontaneous aspermatogenesis was shown not to be a function of differences in the level of testicular testosterone.

Contractility and phenotype transitions in serosal thickening of obstructed rabbit bladder.

Partial outlet obstruction of rabbit bladder induces serosal thickening and smooth muscle (SM) cell hypertrophy that are accompanied by phenotypic changes in the expression of cytoskeletal and cytocontractile proteins. In the present study, we compare the observed progressive phenotypic changes with the contractile responses of strips of the thickened serosa. At 15 days after partial outlet obstruction, although cells in thickened serosa demonstrate the presence of nonmuscle (NM) myosin of A-like type, vimentin, and SM alpha-actin, no contractile responses of this tissue were noted. At later times (30 days), this tissue expressed in addition SM myosin, and this pattern was paralleled by the development of KCl-stimulated contractility. It is only after 60 days that the serosa demonstrated the expression of desmin, phosphoglucomutase (PGM)-related protein, and was locally negative for NM myosin, indicating a maturation toward adult SM cells. Concomitant to this phenotypic change, the response to KCl increased, and a bethanechol-stimulated contractile response developed. At no time period did the serosal layer react with anti-synaptophysin or anti-neurofilament proteins nor did the strips respond to field stimulation (via release of neurotransmitters), showing that SM cell differentiation and development of contractile responses during serosal thickening are independent of innervation.

Genetic and cellular characteristics of bladder outlet obstruction.

Urinary bladder outlet obstruction is a common medical problem. In order to understand the effects of outlet obstruction on bladder morphology, physiology, and pharmacology, several animal models of obstruction have been developed using a variety of species. Although there are marked differences in bladder size, capacity, compliance, physiology, and pharmacology among these species, responses to outlet obstruction have many common characteristics. This article will be separated into six areas: introduction, genetic factors mediating the response during the initial period of partial outlet obstruction and overdistension, cytostructural alterations that accompany compensated bladder function, alterations in innervation accompanying bladder hypertrophy secondary to partial outlet obstruction, alterations in calcium translocation during bladder hypertrophy, and metabolic factors involved in the response to partial outlet obstruction.

Bladder function in experimental outlet obstruction: pharmacologic responses to alterations in innervation, energetics, calcium mobilization, and genetics.

The two functions of the urinary bladder is to store urine at low intravesical pressures, and to periodically expel the urine through a coordinated contraction of the bladder and relaxation of the urethra. To a large extent, urinary bladder function depends upon the underlying structure of the organ as a whole, particularly on the inter-relationships among the smooth muscle, connective tissue, and neuronal elements. An alteration in the ratio of connective tissue to smooth muscle, for example, can significantly alter compliance and functional capacity, structurally impairing the bladder's ability to empty efficiently and fully. Thus, a change in structural compartmentation can affect bladder function independent of autonomic receptor density, response to receptor stimulation, and the contractile capabilities of the smooth muscle elements. Similarly, a selective alteration in either the afferent or efferent innervation of the bladder or urethra can induce significant alterations in the structural interrelationships between smooth muscle and connective elements. In addition, the bladder responds rapidly to alterations in urine volume and urethral resistance with marked changes in bladder and urethral structure and function, and these changes are under the controls of specific genes that are known to control cellular growth, hypertrophy, and hyperplasia. A knowledge of the mechanisms that control the response to specific forms of stress may lead to novel therapies for specific disease states.

Influence of cattle breed and ageing time on textural meat quality.

An experiment was carried out using 7 month old male calves from four different breeds: 10 Spanish Holstein (dairy), 10 Old Brown Swiss (dual purpose), 10 Limousin (fast growth) and 10 Blonde d'Aquitaine (high muscularity). They were all fed concentrate and cereal straw ad libitum. Animals were slaughtered at usual commercial weights for the Spanish market depending on the growth and precocity of each breed (500 kg liveweight at slaughter for Holstein, 550 kg for Old Brown Swiss, 560 kg for Limousin and 620 kg for Blonde d'Aquitaine, with an age between 13 and 15 months). There were significant differences (P<0.001) among breeds in the quantity of total and insoluble collagen in the Longissimus thoracis and lumborum muscle, but collagen solubility was similar (41-44%), except for muscle from the Old Brown Swiss (33%; P<0.001). Overall, breeds specialized for beef production (Limousin and Blonde d'Aquitaine) had lower values for compression and WB in raw and heated meat, respectively, for ageing periods of less than a week. Ageing had a larger effect on myofibrillar tenderness than breed and tended to eliminate the breed effect on textural variables, as well as individual differences within each breed.

Microvasculature of the rabbit urinary bladder.

BACKGROUND: The urinary bladder requires a rich blood supply to maintain its functions, the storage and release of urine. Specialized properties of the bladder vasculature might be anticipated to ensure the integrity of this blood supply, because it is known that blood flow is reduced by distension during bladder filling. However, the bladder vasculature has been described in detail only at the gross level. A comprehensive, three-dimensional view of the blood supply to the bladder wall is presented here. METHODS: The microvasculature of the bladder of male New Zealand white rabbits was described using the combination of vascular corrosion casting, alkali digestion, light microscopy, and scanning and transmission electron microscopy. Following administration of an anticoagulant and an overdose of anesthetic, the abdominal aorta was cannulated just above the inferior mesenteric artery to permit flushing of the distal vasculature. The bladder vasculature was cleared of blood with buffered saline and then either perfuse-fixed with buffered 2% glutaraldehyde and sectioned, or filled with "Mercox" resin to prepare vascular corrosion casts. Casts were cleaned with NaOH, formic acid, and water. In some cases fixed bladders were partially digested with NaOH to expose the mucosal capillary plexus. RESULTS: The bladder is supplied with blood by single, left and right vesicular branches of the internal or external iliac arteries. The serpentine vesicular arteries extend along the lateral borders of the bladder from base to apex just deep to the serosal surface and send dorsal and ventral branches to supply the dorsal and ventral bladder walls. Veins accompany the arteries and exhibit numerous valves. A very dense complex of vessels at the apex of the bladder apparently serves to accommodate bladder distension. The muscularis and submucosa contains few vessels, but the mucosa is well vascularized. An especially dense capillary plexus is present in the lamina propria at its junction with the transitional epithelium. In the relaxed bladder these capillaries lie in grooves formed by the basal layers of the epithelium. The endothelial cells of these capillaries display few cytoplasmic vesicles and are continuous or fenestrated. These capillaries are often invested with pericytes. The mucosal capillary plexus may be associated with an epithelial transport function or may be necessary for urothelial metabolism or maintenance of the barrier function of the urothelium. Unusual capillary tufts, possibly associated with vascular lymphatic tissue, are found associated with the main vessels on the lateral walls in the basal half of the bladder. CONCLUSIONS: These methods present a clear, comprehensive, three-dimensional view of the microvasculature of the bladder wall. They also identify several unique features of this vasculature and provide a basis for studies of the response of this vasculature to pathologic states and experimental manipulation.

Evidence for a unique elastic sheath surrounding the vesicular arteries of the rabbit urinary bladder--studies of the microvasculature with microscopy and vascular corrosion casting.

Because the urinary bladder stores and releases urine, its normal function includes filling and emptying, accompanied by distension and relaxation. It is known that chronic distension compromises blood flow. Recent studies of the rabbit bladder vasculature have described specializations of that vasculature that appear to enhance blood flow in the bladder wall during distension. The present report describes the location, orientation, and structure of an elastic sheath surrounding the vesicular arteries, which may represent one of these specializations. The location, vasculature, and structure of an accessory elastic sheath surrounding the vesicular arteries of the rabbit bladder is described using light and electron microscopy, India ink injection, and vascular corrosion casting. The common iliac arteries of rabbits were cannulated to permit perfusion of the distal vasculature including the urinary bladder. After the bladder vasculature was visually cleared of blood by perfusion with buffered saline, one of the following procedures was used: 1) for light or electron microscopy, the bladder was perfuse-fixed with buffered 2% glutaraldehyde; 2) the bladder vasculature was filled with India ink for vessel tracing; or 3) corrosion casts of the bladder vasculature were prepared by infusion of a Mercox resin mixture. Casts, cleaned of tissue with KOH, and water and formic acid rinses, are dried, and mounted for routine scanning electron microscopy. The presence of an accessory sheath surrounding the main vesicular arteries and some of their branches in the basal two thirds of the urinary bladder was observed on India ink injected specimens and confirmed by micrographs and vascular corrosion casts. The sheath consists of elastic and collagenous fibers and is separated from the tunica media of the arteries by a loose connective tissue layer of variable width. The sheath is circumscribed by a layer of fine blood vessels. The vesicular arteries undulate within the sheath to an extent which is dependent upon the degree of distension of the bladder. This sheath likely represents a specialization which permits the bladder vasculature to accommodate expansion and contraction of the wall during normal filling and emptying. Undulations or coiling of the vesicular arteries within the loose connective tissue core of the sheath increase with bladder contraction, and apparently the sheath simply holds the artery in position during such coiling. The sheath, may represent a modification of the external elastic lamina found in some muscular arteries.

Structure and blood supply of intrinsic lymph nodes in the wall of the rabbit urinary bladder--studies with light microscopy, electron microscopy, and vascular corrosion casting.

The urinary bladder is especially subject to infection by virtue of its direct connection to the external urethral opening, and it is natural to anticipate the presence of a well-developed immunological mechanism to respond to this potential threat. The present study describes small, very highly vascular lymph nodes located in the wall of the rabbit bladder, which may be involved in a local response to foreign antigens. The vasculature and structure of these lymph nodes was described using a combination of vascular corrosion casting, ink injection, and light and electron microscopy. The distal abdominal aorta was cannulated, and after clearing the bladder vasculature with buffered saline, one of the following procedures was used: 1) the bladder was perfuse-fixed in preparation for light and electron microscopy; 2) the bladder vasculature was filled with India ink for vessel tracing; or 3) vascular corrosion casts of the vasculature were prepared by infusing resin comprised of a mixture of Mercox, methyl methacrylate monomer, and catalyst. The resulting casts were cleaned with KOH, formic acid, and water in preparation for scanning electron microscopy. Vascular casts and India ink injections revealed the presence of a number of isolated capillary tufts consisting of clusters of one to five "glomeruli," closely associated with the major vesicular vessels along the lateral walls of the bladder, and supplied by tertiary branches of these vessels. Light and electron microscopy showed that the capillary tufts represented the blood supply to small, ovoid lymph nodes located near the serosal surface of the bladder wall and usually restricted to the basal half of the bladder. These nodes were encapsulated and exhibited subcapsular sinuses, numerous small blood vessels, a limited number of high endothelial cells, and, occasionally, nerves and a follicular substructure. The nodes contained abundant lymphocytes, stellate stromal cells, macrophages, and eosinophils, but lacked the obvious cortical and medullary organization and germinal centers often seen in larger lymph nodes. Vascular corrosion casts, vascular ink injections, and microscopic examination confirmed the presence of small, highly vascular lymph nodes closely associated with the main vesicular vessels along the lateral walls of the rabbit bladder. A follicular substructure of the nodes appears to correspond with the "glomerular" capillary arrangement within the nodes as seen with corrosion casts. The rich blood supply may be indicative of the high metabolic demand of lymphatic tissue, and may be altered in response to the level of activity of the node. The close association between the lymphatic tissue and the rich blood supply to the nodes may allow a rapid mobilization of lymphocytes during a local immune response to foreign agents.

Expression of constitutive heat shock protein-70 in normal (non-stressed) rabbit urinary bladder tissue.

The expression of constitutive HSP-70 in the urinary bladder was determined by SDS-PAGE and western blotting using a mouse monoclonal antibody against HSP-70. The western blot analysis showed that the mouse anti-HSP-70 cross-reacted with a 70 kDa protein present in the extracts of the urinary bladder muscle and mucosa. Densitometric scanning of the western blots allowed us to specifically quantitate the relative amounts of the HSP-70. The quantitation of the HSP-70 by combining immunoblotting and densitometry using a laser scanner is reproducible and this technique requires only a small amount of tissue. The amounts of HSP-70 can be estimated from a standard curve of nanogram(ng) of HSP-70 vs absorption from the immunoblots. The amounts of HSP-70 in the muscular and mucosal layers in the body of the urinary bladder are more than those in the base of the bladder. The presence of HSP-70 in the muscle and mucosal epithelium of the bladder was demonstrated by immunohistochemical analysis of freshly removed tissue from the base and the body of bladder from normal animals.

Rabbit as a model of urinary bladder function.

Micturition is a complex neuromuscular process. Although control mechanisms have been identified at several levels of the central nervous system and spinal cord, the final pathway in the control of micturition is the autonomic innervation of the urinary bladder and related structures. Following this line of reasoning further, micturition is ultimately dependent on the ability of the urinary bladder to both contract and generate intravesical pressure, and to modify its shape in such a way as to efficiently expel its contents without leaving a high residual volume. In order to understand the various elements of micturition, a wide variety of both in vivo and in vitro animal models has been developed. In many cases, animal models have been utilized to describe the effect of specific experimental pathologies on the lower urinary tract. The current review of the use of the rabbit in urological research is not meant to be a comprehensive treatise on the topic, but should provide a rational description of the how this species can be utilized to study both normal and pathological function.

Stimulation of DNA synthesis in rabbit bladder wall after partial outlet obstruction and acute overdistension.

Partial outlet obstruction of the rabbit urethrovesical junction (UVJ) has been used to induce pathology in the urinary bladder characteristic of obstructive damage observed in humans. The purpose of the experiments reported here was to compare the 3H-thymidine (3H-TdR) labelling of DNA in urinary bladders of male New Zealand White (NZW) rabbits subjected to partial outlet obstruction or overdistension. A total of 18 animals was used. Two normal controls, and 12 partially obstructed animals (at 1 day [D], 3D, 5D, 7D, 14D, and 21D) were injected (i.v.) with 3H-TdR at a dose of 0.5 microCi/g body weight. An additional 4 were overdistended to volumes 120% of maximum intravesical pressure, immediately emptied via the catheter, and injected with 3H-TdR 24 hr (1D) later. All animals were sacrificed up to 3.5 hr after injection of the label. DNA-associated radioactivity reached a peak at 3D after obstruction and was reduced substantially by 5D, although the level of incorporation remained well above control levels out to 21D. Levels of 3H-TdR incorporation 1D after overdistension bladders were about half of that found 1D following partial obstruction. The distribution of 3H-TdR labelled DNA in tissues was demonstrated by radioautography of histologic sections. One day following obstruction, 3H-Tdr incorporation was localized in the urothelium. Labelling of urothelium subsequent to 1D was reduced but remained above control levels until 21D. Labelled smooth muscle nuclei were observed only in control and 3D bladders, and they were measured at similar frequencies. Labelling of both intrinsic connective tissue (ICT) (mucosal, submucosal, and mural) and extrinsic connective tissue (ECT) (serosal) peaked at 3D after obstruction and declined thereafter but not to control levels. Labelling of ECT was, of course, limited to those bladders in which ECT was present (i.e., 3-21D). While the distribution of labelled cells in radioautograms was more variable 1D after obstruction than 1D after overdistension, the general cellular and biochemical responses to overdistension, as measured by DNA synthesis, are similar to those observed after partial outlet obstruction. Since the first sequela of obstruction is acute distension, these data support the assertion that the initial overdistension of the bladder initiates the cellular response to obstruction.

Hyperplasia and apoptosis. Opposing cellular processes that regulate the response of the rabbit bladder to transient outlet obstruction.

BACKGROUND: Partial obstruction of the rabbit urethra induces rapid bladder growth. This growth is characterized by hypertrophy of smooth muscle cells in addition to hyperplasia of cells in the urothelium and serosa. The local synthesis of growth factors has been proposed to be influential in this growth since partial outlet obstruction rapidly increases the bladder's expression of basic fibroblast growth factor, while suppressing the expression of transforming growth factor-beta. Upon release of the outlet obstruction, the hypertrophied bladder regresses to its normal weight. Here, we examined whether regression of the hypertrophied rabbit bladder involves apoptosis (programmed cell death) of specific cellular elements and whether the expression of growth factors is altered concomitant with apoptotic cell deletion. EXPERIMENTAL DESIGN: Regressing rabbit bladders were analyzed for markers of apoptosis, including DNA fragmentation and histology. An in situ enzymatic immuno-histochemical procedure was utilized to localize apoptotic cells in these tissues. Finally, Northern blot analysis was used to identify changes in the expression of basic fibroblast growth factor and transforming growth factor-beta during bladder regression. RESULTS: Regressing rabbit bladders demonstrated the characteristic electrophoretic "ladder" pattern of DNA fragmentation associated with apoptosis. An in situ technique to distinguish cells with degraded nuclear DNA identified apoptosis only within the urothelium and serosal lamina of the regressing bladders. RNAs extracted from regressing bladders exhibited decreased expression of basic fibroblast growth factor mRNA as well as increased expression of transforming growth factor-beta 1 mRNA when compared with RNAs from hypertrophied bladders. CONCLUSIONS: We conclude that hyperplasia and apoptosis are opposing cellular processes that mediate the bladder's response to short-term obstructive stimuli and that local synthesis of growth-promoting and growth-inhibitory factors may be responsible for initiating both of these responses.

Indigocarmine as a quantitative indicator of urothelial integrity.

There is increasing evidence that the urothelium of the bladder mucosa prevents the penetration of solutes from the urine into the bladder wall. In the current study, in vivo treatments of rabbit urinary bladder with DMSO, acetone and overdistension resulted in damage to the physical integrity of the bladder mucosa as quantitated by the penetration of the dye indigocarmine (1% in saline) into submucosal tissues. Penetration of the dye can be quantitated, because the dye can be extracted from the tissue and measured spectrophotometrically. Indigocarmine does not penetrate normal, control, bladder mucosae. Bladders treated with gentle 20, 30 and 50% acetone washes for one minute permit dye penetration which is proportional to the acetone concentration utilized. Intravesical 50% DMSO ("RIMSO 50") administration permits modest dye penetration. Distension by slow filling with saline to volumes 90% of capacity and greater causes a marked increase in dye penetration which is proportional to the magnitude of overdistension. Although pretreatment of the bladder with heparin did not reduce the dye penetration following acetone administration, it completely abolished penetration of the dye following overdistension. Indigocarmine is potentially useful as both a quantitative and qualitative indicator of bladder mucosal integrity.

3H-thymidine uptake by the rat urinary bladder after induction of diabetes mellitus.

Streptozotocin-induced diabetes mellitus causes diuresis, increases in bladder mass and changes in micturition. Temporal changes in micturition and bladder mass after induction of diabetes with streptozotocin were monitored and correlated with DNA synthesis and 3H-thymidine incorporation. There were increases in water consumption, urine excretion, urinary frequency, and mean and maximal micturition volume within 1 day after induction of diabetes. These parameters reached maximal values within 6 to 11 days and were maintained at 30 and 60 days. Bladder mass was significantly elevated by 7 days and did not increase further with increasing duration of diabetes. DNA concentration was decreased in bladders from 4, 7 and 14 day diabetics. 3H-thymidine incorporation into DNA increased within 2 days after induction of diabetes, reached maximal values at 4 to 7 days and declined to control values by 14 days. Autoradiography showed intense labelling of the urothelium one day after induction of diabetes, with labelling remaining high up to day 7. Connective tissue and smooth muscle labelling were slower to develop. Labelling of smooth muscle was transient, appearing only on days 4 and 7. The time course of the events was consistent with the hypothesis that bladder distension or increasing micturition volume stimulates thymidine incorporation into DNA, resulting in an increase in bladder mass.

Urothelial permeability of the isolated whole bladder.

The urothelium of the bladder presents an effective barrier to the penetration of solutes from the urine into the bladder wall. Previously, we have demonstrated that the dye indigocarmine can be utilized intravesically to study urothelial permeability. In general, intravesical indigocarmine (administered in vivo) will not penetrate the bladder wall unless the urothelium is damaged by overdistension, acetone administration, or mechanical damage. Unfortunately, using in vivo methodologies, one is limited in the study of the effect of specific conditions and permeations on bladder permeability. In the current study an isolated in vitro whole bladder model was developed to quantitatively study the permeability of the bladder urothelium. In these studies, the penetration of indigocarmine into and through the bladder wall was quantitated under various conditions. The in vitro bladder was filled by infusing 1% indigocarmine in saline in a step-wise manner at the rate of 10 ml in 10 minutes followed by a stabilization period of 10 minutes. Samples were taken from the bath at 20 minutes intervals for spectrophotometrical analysis of the dye. At the end of experiment the bladder was washed in saline for 10 minutes, and stored and extracted in formalin. In general, no indigocarmine penetrated the urothelium until the in vitro capacity was reached and exceeded. At intravesical volumes greater than capacity, the dye concentration in the bath increased very rapidly, even though the integrity of the bladder wall remained intact. In bladders treated with a gentle 50% acetone wash for 1 minute the dye started to penetrate into the bath at intravesical volumes of 25% of capacity and increased rapidly thereafter.(ABSTRACT TRUNCATED AT 250 WORDS)

Pharmacological responses of rabbit urinary bladder after subtotal cystectomy.

Partial cystectomy is used clinically in specific circumstances. There have been some reports indicating that the bladder following subtotal cystectomy can regenerate to a certain degree. The present study investigates the physiology and pharmacology of bladder regeneration at eight weeks after resection of a major part of the bladder body in rabbits. The following studies were performed on control and cystectomy bladders: 1) in vivo cystometry (prior to and four weeks after the operation, and before the sacrifice at eight weeks); 2) sequential muscle strip study (the strips were obtained by dividing bladder transversely into upper body, lower body, mid-bladder, upper base and lower base); and 3) in vitro whole bladder studies. The results can be summarized as follows: 1) grossly there was no regeneration of the bladder body; the cystometric capacity was increased slowly after the operation primarily due to increased size and capacity of the bladder base. 2) The contractile response of the subtotal cystectomized bladder body to field stimulation and bethanechol stimulation was less than the response of normal bladder body. 3) The contractile response to epinephrine showed that the ratio of alpha/beta adrenergic response was much greater in the cystectomy bladder than in the normal bladder. 4) In the whole bladder study, the intravesical pressure response to field stimulation was about the same in both groups, the response to bethanechol was less for cystectomized bladder, and the response to methoxamine was greater for the cystectomized bladder. 5) The ability of cystectomy bladder to empty in response to both field stimulation and bethanechol was impaired whereas the control bladder fully emptied to both field stimulation and bethanechol. In conclusion, over the eight week period following subtotal cystectomy the capacity increased via distention (and hypertrophy) of the bladder base area as opposed to regeneration of the bladder body.

Effects of acute in vitro overdistension of the rabbit urinary bladder on DNA synthesis.

Urinary bladder outflow obstruction induces a myriad of structural and functional changes in the organ. Among the morphological responses to outlet obstruction is both hyperplasia and hypertrophy of specific cellular elements. The initial event which has been implicated in the initiation of the response to outflow obstruction is an initial period of high intravesical pressure and subsequent distention of the bladder. In a previous study, it was shown that at one day following partial outlet obstruction there was a marked increase in thymidine labelling of the urothelium, at 3-5 days, the labeling shifted from the urothelium to the interstitial and serosal elements. The current study was designed to determine if acute distention of the urinary bladder can induce an increase in DNA synthesis (3H-thymidine incorporation), and localize the increased DNA synthesis via autoradiography of 3H-thymidine. In this study, the bladders of adult male New Zealand white rabbits were mounted in isolated in vitro baths. Each control bladder was filled to either 5 or 20 ml. with saline, or distended to 120% of capacity. The bladders were incubated for 7 hours at which time 3H-thymidine was placed both within and outside the bladder for an additional one hour. At the end of the time the bladder was divided at the ureteral orifices into bladder body and base, and each body and base divided into two sections. One section of bladder body and base was quantitatively analyzed for both labelled and unlabelled DNA; the second section was fixed and prepared for autoradiography. The results can be summarized as follows: 1) Acute overdistention for 8 hours induced a slight decrease in the DNA concentration which was mediated by edema of the bladder wall. 2) Acute overdistention induced a 5-fold increase in 3H-thymidine incorporation in the bladder body and a 3-fold increase in the bladder base. Radioautoradiography of the overdistended bladders showed significant and substantial labelling which was confined to the urothelial basal cells. The control bladders showed little or no labelling. These results are consistent with the theory that acute distention following partial outlet obstruction initiates the proliferative response of the bladder to outlet obstruction, and the urothelium is the initial target of the proliferative response. Functionally, the proliferative response may serve to maintain the structural as well as functional integrity of the bladder.

Effect of outlet obstruction on 3H-thymidine uptake: a biochemical and radioautographic study.

Experimental outlet obstruction in the rabbit is characterized by a rapid and substantial increase in urinary bladder mass. Although it is clear that both the smooth muscle and connective tissue compartments are increasing in mass, there is little information on the mechanisms by which this increase in mass occurs. As an initial investigation in this process, urinary bladders from normal and obstructed NZW rabbits were exposed in vitro to tritiated thymidine (3H-TdR) in order to determine which populations of cells are induced to synthesize DNA following outlet obstruction, and when, after obstruction, such synthesis occurs. Biochemical analysis of nucleic acids was performed on each specimen to determine total and radioactive DNA. These analyses showed a marked increase in DNA synthesis at 24 hours following obstruction which remained relatively high through seven days after obstruction. There was a decline in labelling at 14 days. Incorporation of radioactive label peaked at three days and declined to control levels by 14 days. Samples of tissue were taken from each subject and processed for radioautography. At 24 hours after obstruction, significant numbers of cells of the basal cell layer of the urothelium are observed to be actively involved in DNA synthesis, while the other two tissue compartments (muscularis and connective tissue) show no significant changes when compared to normal specimens. Connective tissue, on the other hand, showed significantly increased levels of labelling above control level from three to 14 days after obstruction. Smooth muscle cells were observed to be frequently labelled in only one of the experimental bladders observed three days after obstruction.